|Dynamics of Host-Pathogen Interactions|
|HEAD||Jost Enninga, Ph.D / firstname.lastname@example.org|
|MEMBERS||Mr. Laurent Audry (technician, Institut Pasteur), Mr. Alexandre Bobard, Ph.D. (postdoc), Melle Soudeh Ehsani (Ph.D. student, University of Paris VII), Mme Anne-Marie Fargues (administrative assistant), Melle Geneviève Janvier (engineer, Institut Pasteur), Melle Cristina Dias Rodrigues, Ph.D. (postdoc)
We are interested in deciphering the molecular and cellular mechanisms of bacterial invasion of host cells. Therefore, our laboratory connects the fields of cell biology, microbial pathogenesis, and immunology by developing novel, fluorescent microscopic approaches. Our aim is to establish approaches that correlate the pathogen behaviour and the host responses in single experiments at cellular and subcellular resolution. We typically use Shigella flexneri, the causative of bacterial dysentery, as model pathogen for assay development. Currently, we are following three major axes of research as outlined below.
1) Injection of bacterial effectors into host cells (Cristina Dias Rodrigues, Laurent Audry and Jost Enninga)
One strategy employed by a number of pathogens is the secretion of bacterial effector proteins into host cells via sophisticated secretion machinery. The type III secretion system is one such machine resembling a molecular syringe. A particularity of the type III secretion needles is their capacity to translocate bacterial effectors in a single step across three membrane layers- the inner and outer bacteria membranes, and the host plasma membrane. We have set up a novel methodology for direct fluorescent labelling of bacterial effectors without functional loss within living bacteria. Using rapid multidimensional fluorescence microscopy, we can track the pace of effector secretion from individual bacteria. This enables the comparison of the secretion kinetics of the translocated effectors, we can track where secretion takes place, and we can correlate the effector secretion with the host responses induced.
2) Localization of invasive bacteria inside host cells (Alexandre Bobard and Jost Enninga)
Cellular uptake of bacterial pathogens leads to (i) blocking the progression of the bacteria-containing endosomal vacuoles and their subsequent fusion with lysosomes (e.g. M. tuberculosis), (ii) altering the composition of the endosomal vacuoles that appear advantageous to the invading pathogen (e.g. Salmonella), or (iii) escaping the endocytic vacuole via rupture of the enclosing membrane (e.g. Shigella). Studying these events has given novel insight into the mode of function of the participating constituents of the vesicular trafficking machine. However, the overall sequence and hierarchy of the involved steps are still poorly understood due to the limited number of approaches that allow dynamic and functional investigations at the single cell level. We are establishing novel fluorescence microscopy based approaches that give precise information on the intracellular localization of bacterial pathogens with very high temporal resolution. With our novel assays, we are able to study how bacterial factors communicate with host factors to shape the localization of the pathogens during cellular uptake.
3) Host immune responses and how they are undermined (Soudeh Ehsani, Geneviève Janvier and Jost Enninga)
During the internalization of bacterial pathogens, a number of immune genes are induced. The expression of host genes depends on the behaviour of the pathogens and on their intracellular localization. Common reporters for gene expression, like GFP or luciferase, have the disadvantage that they require maturation- a process that takes time which is difficult to predict. Furthermore, other approaches for gene profiling, like gene chips require the disruption of the cells and collection of transcripts from a large number of cells. To overcome these problems we have developed novel reporter systems that allows gene profiling of specific immune genes in single living cells. We are able to combine our novel approaches with assays that track the localization of invading bacteria and their physiological behaviour. This leads to integrative information on the invasion strategies employed by the pathogens and on the host responses induced.
Enninga J, Mounier J, Sansonetti P, Tran Van Nhieu G. 2005. Studying the Secretion of Type III Effectors into Host Cells in Real Time.Nature Methods. Dec;2(12):959-65.
Pulliainen AT, Enninga J, Fernandez-Arenas E, Griffiths G. A course with a difference. 2007. Fighting infectious diseases with technology and knowledge-transfer.EMBO Rep. Feb;8(2):117-20.
Cabal G., Enninga J., Mhlanga MM. 2007. Single Molecule Tracking inside Living Samples.Bookchapter in “Imaging Cellular and Molecular Biological Function” (editors: Shorte SL and Frischknecht F). Springer Publishers: 235-263
Enninga J., Sansonetti P., TournebizeR. 2007. Roundtrip explorations of bacterial infection: from single cells to the entire host. Trends Microbiol.Nov;15(11):483-90.
Simpson N., Audry L., and Enninga J. 2008.Tracking the secretion of fluorescently-labeled type III effectors from single bacteria in real time. Methods in Mol Biology. in press
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Activity Reports 2009 - Institut Pasteur
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